title: Functional and physical molecular size of the chicken hepatic lectin determined by radiation inactivation and sedimentation equilibrium analysis.
abstract: Radiation inactivation and sedimentation equilibrium analysis were used to determine the functional and physical size of the chicken hepatic membrane receptor that binds N-acetylglucosamine-terminated glycoproteins. Purified plasma membranes from chicken liver were irradiated with high energy electrons and assayed for 125I-agalactoorosomucoid binding. Increasing the dose of ionizing radiation resulted in a monoexponential decay in binding activity due to a progressive loss of binding sites. The molecular mass of the chicken lectin, determined in situ by target analysis, was 69,000 +/- 9,000 Da. When the same irradiated membranes were solubilized in Brij 58 and assayed, the binding protein exhibited a target size of 62,000 +/- 4,000 Da; in Triton X-100, the functional size of the receptor was 85,000 +/- 10,000 Da. Sedimentation equilibrium measurements of the purified binding protein yielded a lower limit molecular weight of 79,000 +/- 7,000. However, the solubilized lectin was detected as a heterogeneous population of oligomers with molecular weights as high as 450,000. Addition of calcium or calcium plus N-acetylglucosamine decreased the higher molecular weight species, but the lower limit molecular weights remained invariant. Similar results were determined when the chicken lectin was solubilized in Brij 58, C12E9, or 3-[(3-cholamidopropyl)dimethylammonio]-1-propane-sulfonic acid (CHAPS). Results from the present study suggest that in the plasma membrane, the functional species of the chicken hepatic lectin exists as a trimer. However, in detergent solution, the purified receptor forms a heterogeneous population of irreversible oligomers that exhibit binding activity proportional to size.
authors.affiliation: Laboratory of Biochemistry and Metabolism, National Institute of Diabetes, Digestive and Kidney Diseases, National Institutes of Health, Bethesda, Maryland 20892.
authors.forename: C J
authors.initials: CJ
authors.lastname: Steer
authors.forename: J C
authors.initials: JC
authors.lastname: Osborne
authors.forename: E S
authors.initials: ES
authors.lastname: Kempner
mesh_terms: D000818:Animals; D037263:Asialoglycoprotein Receptor; D002458:Cell Fractionation; D002462:Cell Membrane / Q000378:metabolism / Q000648:ultrastructure; D002645:Chickens; D008099:Liver / Q000378:metabolism / Q000528:radiation effects; D046911:Macromolecular Substances; D008970:Molecular Weight; D011971:Receptors, Immunologic / Q000302:isolation & purification / Q000378:metabolism* / Q000528:radiation effects
chemical_list: D037263:Asialoglycoprotein Receptor; D046911:Macromolecular Substances; D011971:Receptors, Immunologic